Protein-protein interactions underlying molecular recognition are studied using monoclonal antibodies directed against the model protein antigen, lysozyme c (HEL). The X-ray structures of two Fab-HEL complexes for 2 of these antibodies (HH5 and HH10) have been refined; we are now studying in detail the molecular mechanisms underlying specificity and affinity in protein-protein interactions. A panel of hybridomas representing various time points in the immune response to HEL was functionally characterized by several different types PCFIA. Of over 500 primary and secondary response hybridomas analyzed, no IgM antibodies were specific for HEL, although HEL-specific IgM precursor cells are available. There is no obvious pattern of affinity "maturation" during the course of the IgG response. Most specificities are found throughout the response, and no single specificity dominates any one time point, although a few specificities are confined to very late in the immune response. All the antibodies analyzed to date fall into one of 4 complementation groups, three of which have been described previously and include the majority of the antibodies; antibodies within a complementation reciprocally cobind (complement) and competitively block one another. The patterns of interaction support the hypothesis that the entire surface of the protein is antigenic. They also suggest a fundamental relationship between protein structure and antigenicity. Analyses of interactions among three antibodies whose X-ray crystallographic structures are known suggest the apparent structural overlap of epitopes will not necessarily result in competitive exclusion, and that those antibodies showing strong competitive interference overlap significantly in their contacts with HEL. Site-directed mutations both of expressed antibody and HEL genes are being utilized to examine the specific roles of individual amino and residues to molecular specificity.